Systematical mutational analysis of teriparatide on anti-osteoporosis activity by alanine scanning.

Osteoporosis is a progressive systemic skeletal disease that decreases bone density and bone quality, making them fragile and easy to break. In spite of effective anti-osteoporosis potency, teriparatide, the first anabolic medications approved for the treatment of osteoporosis, was proven to exhibit various side effects. And the relevant structure-activity relationship (SAR) of teriparatide was in need. In this work, we performed a systematical alanine scanning against teriparatide and synthesized 34 teriparatide derivatives. Their biological activities were evaluated and the importance of each residue for anti-osteoporosis activity was also revealed. A remarkable decrease in activity was observed for alanine replacement of the residue Gly12, His14, Ser17, Arg20 and Leu24, showcasing the important role of these residues in teriparatide on anti-osteoporosis activity. On contrary, when Gly13 and Gln30 were mutated to Ala, the peptide derivatives exhibited the significantly increased activities, demonstrating that these two residues could be readily replaced. Our research expanded the peptide library of teriparatide analogues and presented a potential opportunity for designing the more powerful anti-osteoporosis peptide agents.

In vivo dynamic analysis of BMP-2-induced ectopic bone formation.

Bone morphogenetic protein (BMP)-2 plays a central role in bone-tissue engineering because of its potent bone-induction ability. However, the process of BMP-induced bone formation in vivo remains poorly elucidated. Here, we aimed to establish a method for intravital imaging of the entire process of BMP-2-induced ectopic bone formation. Using multicolor intravital imaging in transgenic mice, we visualized the spatiotemporal process of bone induction, including appearance and motility of osteoblasts and osteoclasts, angiogenesis, collagen-fiber formation, and bone-mineral deposition. Furthermore, we investigated how PTH1-34 affects BMP-2-induced bone formation, which revealed that PTH1-34 administration accelerated differentiation and increased the motility of osteoblasts, whereas it decreased morphological changes in osteoclasts. This is the first report on visualization of the entire process of BMP-2-induced bone formation using intravital imaging techniques, which, we believe, will contribute to our understanding of ectopic bone formation and provide new parameters for evaluating bone-forming activity.

Synergistic effect of hyperbaric oxygen therapy with PTH [1-34] on calvarial bone graft in irradiated rat.

OBJECTIVE: To determine the synergistic effect of parathyroid hormone (PTH) [1-34] in combination with hyperbaric oxygen (HBO) on bone graft in a rat calvarial bone defect model under impaired osteogenic conditions. MATERIALS AND METHODS: Twenty-four rats were divided into three groups. Localized radiation with a single 12 Gy dose was administered to the calvaria. Four weeks after radiation, calvarial circular defects were created in the parietal bones. All defects were filled with biphasic calcium phosphate. After the bone graft, PTH [1-34] was injected subcutaneously, and HBO was administered. At 6 weeks after the bone graft, the rats were sacrificed, and specimens were harvested. RESULTS: Histomorphometric evaluation showed that the percentage of new bone area was higher in the PTH and PTH/HBO groups than in the control group. The percent residual material area was decreased in the PTH/HBO group compared with the control group. The percentage blood vessel number was highest in the PTH group. Micro-CT evaluation showed that the new bone volume was highest in the PTH/HBO group. The residual material volume was lowest in the PTH/HBO group. CONCLUSION: Within the limitations of this study, our data indicate that PTH combined with HBO may reverse radiation-induced impairment of bone healing.

Calcium-Triggered Pulsatile Delivery of Parathyroid Hormone from Microbeads for Osteoporosis Treatment.

Recombinant human parathyroid hormone 1-34 (rhPTH 1-34) is the most potent anabolic drug recommended for patients with osteoporosis who do not respond to conventional treatment. However, subcutaneous intermittent injection is the only effective regimen due to its unusual action of mechanism. This regimen is inconvenient and is a big hurdle in clinical applications. In this study, we designed polyelectrolyte microbeads that can deliver rhPTH 1-34 in response to Ca2+ concentration, which indicates the osteoporotic status. Dextran photopolymer was synthesized, mixed with anionic monoacrylate, and photopolymerized by passing through capillary microfluidics to obtain the microbeads. The anionic property of microbeads was confirmed by toluidine blue staining. One microbead, loaded with a 1 day dose of rhPTH 1-34 (23.4 ± 0.9 µg), released rhPTH 1-34 in a triggered manner following the addition of Ca2+ ion. In vitro cell study demonstrated that rhPTH 1-34 released in a pulsatile manner from the microbeads induced osteogenic markers (ALP, RUNX2, and OPN) and precipitated mineral disposition more effectively.

Nanotechnology Treatment Options for Osteoporosis and Its Corresponding Consequences.

Unfortunately, osteoporosis, as a worldwide disease, is challenging human health with treatment only available for the symptoms of osteoporosis without managing the disease itself. Osteoporosis can be linked as the common cause of fractures and increased mortality among post-menopausal women, men, and the elderly. Regrettably, due to osteoporosis, incidents of fractures are more frequent among the presented populations and can be afflictive for carrying out everyday life activities. Current treatments of osteoporosis encompass changing lifestyles, taking orthopedic drugs, and invasive surgeries. However, these treatment options are not long lasting and can lead to complications after post-surgical life. Therefore, to solve this impairment, researchers have turned to nanotechnologies and nanomaterials to create innovative and alternative treatments associated with the consequences of osteoporosis. This review article provides an introduction to osteoporotic compression vertebral fractures (OVCFs) and current clinical treatments, along with the rationale and efficacy of utilizing nanomaterials to modify and improve biomaterials or instruments. The methods of applying bioactive agents (bone morphogenetic protein-2 (BMP-2), parathyroid hormone 1-34 (PTH 1-34)), as well as 3D printing will be presented from an osteoporosis treatment perspective. Additionally, the application of nanoparticles and nanotube arrays onto the current surgical treatments and orthopedic drug administration methods addressed will show that these systems reinforce a better mechanical performance and provide precise and slow-releasing drug delivery for better osseointegration, bone regeneration, and bone strength. In summary, nanomaterials can be seen as an alternative and more effective treatment for individuals with osteoporosis.

Comparative study of the effect of PTH (1-84) and strontium ranelate in an experimental model of atrophic nonunion.

UNLABELLED: This study aimed to set up an experimental model of long bone atrophic nonunion and to explore the potential role of PTH-1-84 (PTH 1-84) and strontium ranelate (SrR). A model of atrophic nonunion was created in Sprague-Dawley rats at the femoral midshaft level. The animals were randomised into four groups. Group A1: control rodents, fracture without bone gap; Group A2: rodents with subtraction osteotomy (non-union model control) treated with saline; Group B: rodents with subtraction osteotomy treated with human-PTH (PTH 1-84); and Group C: rodents with subtraction osteotomy treated with strontium ranelate (SrR). The groups were followed for 12 weeks. X-rays were be obtained at weeks 1, 6 and 12. After sacrificing the animals, we proceeded to the biomechanical study and four point bending tests to evaluate the resistance of the callus and histological study. In second phase, the expression of genes related to osteoblast function was analysed by reverse transcription-quantitative PCR in rats subjected to substraction osteotomy and treated for 2 weeks. The animals were randomised into three groups: Group A2: rodents treated with saline; Group B: rodents treated with PTH 1-84 and Group C: rodents treated with SrR. RESULTS: No significant histological differences were found between animals subjected to subtraction osteotomy and treated with either saline or PTH (p=0.628), but significant difference existed between animals receiving saline or SrR (p=0.005). There were no significant differences in X-ray score between the saline and PTH groups at either 6 or 12 weeks (p=0.33 and 0.36, respectively). On the other hand, better X-ray scores were found in the SrR group (p=0.047 and 0.006 in comparison with saline, at 6 and 12 weeks, respectively). In line with this, biomechanical tests revealed improved results in the SrR group. Gene expression analysis revealed a slightly decreased levels of DKK1, a Wnt pathway inhibitor, in rats treated with SrR. CONCLUSIONS: SrR increases has a beneficial effect in this atrophic non-union model in rats. This suggests that it might have a role may have important implications for the potential clinical role in the treatment of fracture nonunion.

[Medication and bone metabolism: Clinical importance for fracture treatment]. / Medikamente und Knochenstoffwechsel : Klinische Bedeutung für die Frakturbehandlung.

INTRODUCTION: The improvement and acceleration of fracture healing has been a component of medical practice since fractures have been treated. The aim is not only to fulfill the basic principles of fracture healing, such as reduction, retention, soft tissue coverage and infection prevention but also to reduce negative influences on fracture healing and promote positive factors. Nicotine, alcohol, diabetes and malnutrition can negatively affect fracture healing and should be appropriately controlled during fracture treatment; however, it is far more difficult to develop medicinal treatment strategies that lead to improvement and acceleration of fracture healing. AIM: This article provides an overview of pharmacological factors influencing fracture healing. In addition, substances frequently used in clinical practice will be evaluated in terms of the effects on fracture healing processes. MATERIAL AND METHODS: An extensive literature search was conducted in PubMed based on thematic keywords. The selection of studies and scientific publications focused mainly on results from clinical trials in order to provide practically relevant information. RESULTS: In this context, preclinical studies have identified several drugs that lead to the acceleration of fracture healing; however, only a very limited number of clinical trials have confirmed this positive effect. Most of these studies dealt with drugs developed for the treatment of osteoporosis, as osteoporotic fractures are common and a positive or negative influence of such drugs are of particular interest in this field. In the field of osteoporosis medication a certain degree of positive effect of parathyroid hormone 1-34 (PTH) on fracture healing has been shown in clinical trials. For other osteoporosis medications no negative influence on fracture healing in clinical settings has been reported; however, there seems to be a positive effect in terms of better implant fixation for patients receiving oral bisphosphonate therapy. DISCUSSION: Systemic medication to improve fracture healing will not be part of the clinical routine in the foreseeable future as the available data for already approved drugs and drugs under development do not currently justify routine administration. However, the currently known data should encourage the potential of known medications to be completely exhausted in fracture healing studies as well as novel therapy options in the sense of positive effects on fracture healing in order to improve patient care.

Successful treatment of 1-34 parathyroid hormone (PTH) after failure of bisphosphonate therapy in a complex case of pregnancy associated osteoporosis and multiple fractures.

INTRODUCTION: Pregnancy associated osteoporosis (PAO) was first reported almost half a century ago. The most common symptom is acute lower back pain due to vertebral fractures in the last trimester or immediately after birth. PATIENT: We present a case involving a female patient born in 1971 (gravida II, para I) with a history of PAO. In April 2000 at the age of 28 years, she delivered a son and breastfed him for 4 months. A first magnetic resonance tomography (MRT) screening in June 2000 showed osteoporotic fractures at lumbar vertebra 1-4. Therefore, the patient received oral alendronate therapy. In May 2001, a second MRT exhibited burst fracture of thoracic 8, end-plate fracture of thoracic 11, 12, lumbar 2-5 and compression fracture of lumbar 1. The oral therapy was switched to ibandronate (3 mg) intravenously every 3 months. An X-ray in December 2002 showed 3 new additional end-plate fractures at thoracic 4, 6 and 7. Ibandronate was discontinued in September 2004 and the patient received daily subcutaneous (s. c.) injections of 1-34 PTH in September 2005. RESULTS: After starting 1-34 PTH treatment for 18 months, a further increase in bone mineral density (BMD) was achieved without any further fracture. CONCLUSION: We presented for the first time a case of severe PAO with 11 spine fractures. We observed an unsatisfactory effect of oral and i. v. bisphosphonates in combination with adequate calcium and vitamin D supplementation. The treatment with 1-34 PTH showed an increase in BMD with no further fractures.

Intrinsic bioconjugation for site-specific protein PEGylation at N-terminal serine.

Recently developed chemical ligation protocols were elaborated for rapid N-terminal protein PEGylation. We introduce a PEG-salicylaldehyde ester and demonstrate its site-specific ligation to the N-terminus of the RNase S protein and to the therapeutic polypeptide PTH (1-34).

Semisynthesis of peptoid-protein hybrids by chemical ligation at serine.

Chemical ligation protocols were explored for generating semisynthetic peptoid-protein hybrid architectures containing a native serine residue at the ligation site. Peptoid oligomers bearing C-terminal salicylaldehyde esters were synthesized and ligated to the N-terminus of the RNase S protein or the therapeutic hormone PTH(1-34) polypeptide. This technique will expand the repertoire of strategies to enable design of hybrid macromolecules with novel structures and functions not accessible to fully biosynthesized proteins.

Limited utility of tartrate-resistant acid phosphatase isoform 5b in assessing response to therapy in osteoporosis.

BACKGROUND: Tartrate-resistant acid phosphatase isoform 5b (TRACP5b) is a serum bone resorption marker. Our aim was to investigate its utility in monitoring metabolic bone disease. METHODS: Serum TRACP5b, C-terminal cross-linking telopeptide of type I collagen, urine N-terminal cross-linking telopeptide of type I collagen and free deoxypyridinoline were measured pre- and post-treatment with a parathyroid hormone analogue [PTH (1-34)] (n = 14) or a bisphosphonate (N-BP) (n = 8). TRACP5b, bone alkaline phosphatase (bone ALP), 25-hydroxyvitamin D (25OHD) and parathyroid hormone (PTH) were measured in 100 osteoporosis patients on prolonged bisphosphonate therapy. RESULTS: Changes in TRACP5b were smaller in magnitude but mimicked those of other bone resorption markers. Absolute changes in TRACP5b and the other resorption markers correlated significantly (p < 0.001). In patients on long-term bisphosphonates, TRACP5b and bone ALP levels were not suppressed. Vitamin D status was consistent with the level of supplementation. CONCLUSION: TRACP5b has limited utility as a single marker of metabolic bone disease treatment.

Time-resolved fluorescence ligand binding for G protein-coupled receptors.

G protein-coupled receptors (GPCRs) and their ligands are traditionally characterized by radioligand-binding experiments. These experiments yield excellent quantitative data, but have low temporal and spatial resolution. In addition, the use of radioligands presents safety concerns. Here we provide a general procedure for an alternative approach with high temporal and spatial resolution, based on Tb(+)-labeled fluorescent receptor ligands and time-resolved fluorescence resonance energy transfer (TR-FRET). This protocol and its design are detailed here for the parathyroid hormone receptor, a class B GPCR, and its fluorescently labeled 34-amino acid peptide ligand, but it can be easily modified for other receptors and their appropriately labeled ligands. We discuss three protocol options that use Tb(+)-labeled fluorescent ligands: a time-resolved fluorescence separation option that works on native receptors but requires separation of bound and unbound ligand; a TR-FRET option using SNAP-tag-labeled receptors for high-throughput screening; and a TR-FRET option that uses fluorescently labeled antibodies directed against an epitope engineered into the Flag-labeled receptors' N terminus. These protocol options can be used as standard procedures with very high signal-to-background ratios in order to characterize ligands and their receptors in living cells and in cell membranes via straightforward plate-reader measurements.

ß-arrestin-selective G protein-coupled receptor agonists engender unique biological efficacy in vivo.

Biased G protein-coupled receptor agonists are orthosteric ligands that possess pathway-selective efficacy, activating or inhibiting only a subset of the signaling repertoire of their cognate receptors. In vitro, D-Trp(12),Tyr(34)-bPTH(7-34) [bPTH(7-34)], a biased agonist for the type 1 PTH receptor, antagonizes receptor-G protein coupling but activates arrestin-dependent signaling. In vivo, both bPTH(7-34) and the conventional agonist hPTH(1-34) stimulate anabolic bone formation. To understand how two PTH receptor ligands with markedly different in vitro efficacy could elicit similar in vivo responses, we analyzed transcriptional profiles from calvarial bone of mice treated for 8 wk with vehicle, bPTH(7-34) or hPTH(1-34). Treatment of wild-type mice with bPTH(7-34) primarily affected pathways that promote expansion of the osteoblast pool, notably cell cycle regulation, cell survival, and migration. These responses were absent in ß-arrestin2-null mice, identifying them as downstream targets of ß-arrestin2-mediated signaling. In contrast, hPTH(1-34) primarily affected pathways classically associated with enhanced bone formation, including collagen synthesis and matrix mineralization. hPTH(1-34) actions were less dependent on ß-arrestin2, as might be expected of a ligand capable of G protein activation. In vitro, bPTH(7-34) slowed the rate of preosteoblast proliferation, enhanced osteoblast survival when exposed to an apoptotic stimulus, and stimulated cell migration in wild-type, but not ß-arrestin2-null, calvarial osteoblasts. These results suggest that bPTH(7-34) and hPTH(1-34) affect bone mass in vivo through predominantly separate genomic mechanisms created by largely distinct receptor-signaling networks and demonstrate that functional selectivity can be exploited to change the quality of G protein-coupled receptor efficacy.

Prevention of atrophic nonunion by the systemic administration of parathyroid hormone (PTH 1-34) in an experimental animal model.

OBJECTIVES: Recombinant human parathyroid hormone (PTH 1-34) has been previously shown to enhance fracture healing in animal models. Here, we sought to determine whether the systemic administration of PTH 1-34 is effective in preventing atrophic fracture nonunion in a murine, surgical nonunion model. METHODS: We used an established reproducible long-bone murine fracture nonunion model by generating a midshaft femur fracture, followed by fracture distraction using an intramedullary pin and custom metallic clip to maintain a fracture gap of 1.7 mm. Mice were randomized to receive either daily intraperitoneal injections of 30 µg/kg PTH 1-34 for 14 days or saline injections. At 6 weeks after the procedure, radiographic and histologic assessment of fracture healing was performed. RESULTS: At 6 weeks after surgery, the group treated with PTH showed higher rates of bony union (50% vs 8%; P < 0.05) as assessed by radiographic analysis. Mean gap size was also significantly lower in the PTH group (1.42 vs 0.36 mm in the control group; P < 0.05). Histologic analysis of atrophic nonunions in the control group revealed a persistent fracture gap with intervening fibrous tissue. In contrast, healed subjects in the PTH-treated group had cortical bridging with mature bone and relatively little callus, which is consistent with primary intramembranous ossification. CONCLUSIONS: Daily systemic administration of recombinant PTH 1-34 increased the rate of union in a mouse atrophic nonunion model. This may have important implications for the potential clinical role of PTH 1-34 in the treatment of atrophic fracture nonunions.

The influence of parathyroid hormone treatment on implant fixation.

INTRODUCTION: Primary joint replacements generally function well with excellent clinical results. However, failure rates for young patients are still high and increasing in number. The longterm survival of an uncemented prosthesis is influenced by multiple factors depending on host physiology as well as properties of implanted material, initial mechanical stability, early osseointegration, and the surrounding bone. Parathyroid hormone is the principal regulator of calcium homeostasis and involved in the control of bone remodelling. Parathyroid hormone administered intermittently increases bone formation and mass by osteoblast stimulation. Early osseointegration and implant fixation could potentially be enhanced with adjuvant parathyroid hormone treatment. The aim of the studies in this PhD thesis was to determine if implant fixation of experimental implants can be improved with adjuvant intermittent administration of parathyroid hormone. STUDIES: All studies used an experimental canine model of early implant fixation inserting porous coated titanium alloy implants with no weight bearing in a bed of cancellous bone. The study design was un-paired. Test animals were randomised to PTH (1-34) 5 µm/kg daily for 4 weeks. Implant fixation was defined by mechanical stability and osseointegration. Study I investigated the effect of parathyroid hormone on implant fixation of implants inserted press fit with surrounding bone in the proximal tibia of 20 canines. Histomorphometric analysis showed increased amount of new bone in contact with the implant. No improvement was observed in the surrounding bone. PTH did not increase mechanical fixation in pushout test. Study II investigated the effect of parathyroid hormone on implant fixation of implants surrounded by a critical 1 mm gap. Implants where inserted in the tibia of 20 canines. Bone density was increased in the inner gap and outer gap with PTH treatment. Bone at implant interface improved with PTH but did not achieve significance. Push-out testing showed that PTH Increased mechanical implant fixation in shear stiffness and total energy absorption. Shear strength was not significantly increased. Study III investigated the effect on implant fixation of implants surrounded by a 2.5 mm gap in which morsellised allograft was impacted. Implants were inserted in 20 Canines in the humerus. Histomorphometric analysis showed that PTH increased the amount of new bone within the gap, but not in contact the implant. There were no differences in amount of allograft. The push-out testing showed no differences in mechanical parameters. CONCLUSION: The studies in this PhD thesis demonstrated that parathyroid hormone increases bone healing around implants in situations of insertion in press-fit or in more challenging environments of empty and grafted gaps. Early fixation was increased in implants with gaps, in which pure gap bone stimulation improved fixation. This warrants further preclinical studies.

Anabolic and catabolic regimens of human parathyroid hormone 1-34 elicit bone- and envelope-specific attenuation of skeletal effects in Sost-deficient mice.

PTH is a potent calcium-regulating factor that has skeletal anabolic effects when administered intermittently or catabolic effects when maintained at consistently high levels. Bone cells express PTH receptors, but the cellular responses to PTH in bone are incompletely understood. Wnt signaling has recently been implicated in the osteo-anabolic response to the hormone. Specifically, the Sost gene, a major antagonist of Wnt signaling, is down-regulated by PTH exposure. We investigated this mechanism by treating Sost-deficient mice and their wild-type littermates with anabolic and catabolic regimens of PTH and measuring the skeletal responses. Male Sost(+/+) and Sost(-/-) mice were injected daily with human PTH 1-34 (0, 30, or 90 µg/kg) for 6 wk. Female Sost(+/+) and Sost(-/-) mice were continuously infused with vehicle or high-dose PTH (40 µg/kg · d) for 3 wk. Dual energy x-ray absorptiometry-derived measures of intermittent PTH (iPTH)-induced bone gain were impaired in Sost(-/-) mice. Further probing revealed normal or enhanced iPTH-induced cortical bone formation rates but concomitant increases in cortical porosity among Sost(-/-) mice. Distal femur trabecular bone was highly responsive to iPTH in Sost(-/-) mice. Continuous PTH (cPTH) infusion resulted in equal bone loss in Sost(+/+) and Sost(-/-) mice as measured by dual energy x-ray absorptiometry. However, distal femur trabecular bone, but not lumbar spine trabecular bone, was spared the bone-wasting effects of cPTH in Sost(-/-) mice. These results suggest that changes in Sost expression are not required for iPTH-induced anabolism. iPTH-induced resorption of cortical bone might be overstimulated in Sost-deficient environments. Furthermore, Sost deletion protects some trabecular compartments, but not cortical compartments, from bone loss induced by high-dose PTH infusion.

Effects of intermittent parathyroid hormone treatment on osteoprogenitor cells in postmenopausal women.

Intermittent parathyroid hormone (PTH) 1-34 treatment stimulates bone formation, but the molecular mechanisms mediating this effect have not been previously studied in humans. Thus, we used magnetic activated cell sorting to isolate hematopoietic lineage negative (lin-)/alkaline phosphatase positive (AP+) osteoprogenitor cells from bone marrow of 20 postmenopausal women treated with PTH (1-34) for 14 days and 19 control subjects. Serum PINP and CTX increased in PTH-treated subjects (by 97% and 30%, respectively, P<0.001). Bone marrow lin-/AP+ cells from PTH-treated subjects showed an increase in the RANKL/OPG mRNA ratio (by 7.5-fold, P=0.011) and in the mRNAs for c-fos (a known PTH-responsive gene, by 42%, P=0.035) and VEGF-C (by 57%, P=0.046). Gene Set Enrichment Analysis (GSEA, testing for changes in pre-specified pathways) demonstrated that PTH had no effect on osteoblast proliferation, apoptosis, or differentiation markers. However, PTH treatment resulted in a significant decrease (GSEA P-value, 0.005) in a panel of BMP target genes in the lin-/AP+ cells. Our findings thus identify several future directions for studying mechanisms of PTH action in humans. First, given the increasing evidence that PTH induces angiogenesis, the role of increased VEGF-C production by bone marrow osteoprogenitor cells in mediating this effect and the anabolic response to PTH warrants further study. Second, while the observed inhibition of BMP target gene expression by PTH is not consistent with the anabolic effects of PTH on bone and requires further validation, these data do generate the hypothesis that an inhibition of BMP signaling by PTH may, over time, limit the availability of mature osteoblasts on bone surfaces and thereby contribute to the observed waning of the anabolic response to PTH.

Electrogenic Na⁺/HCO3⁻ co-transporter-1 is essential for the parathyroid hormone-stimulated intestinal HCO3⁻ secretion.

Parathyroid hormone (PTH) was recently demonstrated to enhance the HCO(3)(-) secretion through the apical anion channel, cystic fibrosis transmembrane conductance regulator (CFTR), but how the HCO(3)(-) entered the epithelial cells was not well understood, in part, due to the lack of specific inhibitors of the basolateral HCO(3)(-) transporters. Moreover, the function of the PTH-stimulated HCO(3)(-) secretion has never been investigated in vivo. Here, we designed three specific pairs of small interfering RNA sequences to simultaneously knockdown three variants of the electrogenic Na(+)/HCO(3)(-) co-transporter (NBCe)-1 in the intestinal epithelium-like Caco-2 monolayer. The results showed that NBCe1 mRNA levels were markedly reduced, and the PTH-induced transepithelial current and voltage changes were diminished after triple knockdown as determined by quantitative real-time PCR and Ussing chamber technique, respectively. An in vivo ligated intestinal loop study further showed that there was an increased fluid secretion, presumably driven by HCO(3)(-) transport, in the ileum, but not in jejunum or colon, of rats administered intravenously with 2 µg/kg body weight of rat PTH 1-34. Therefore, the present results suggested that PTH stimulated intestinal HCO(3)(-) secretion, particularly in the ileum, by inducing the basolateral HCO(3)(-) uptake via NBCe1.

Anabolic agents and bone quality.

BACKGROUND: The definition of bone quality is evolving particularly from the perspective of anabolic agents that can enhance not only bone mineral density but also bone microarchitecture, composition, morphology, amount of microdamage, and remodeling dynamics. QUESTIONS/PURPOSES: This review summarizes the molecular pathways and physiologic effects of current and potential anabolic drugs. METHODS: From a MEDLINE search (1996-2010), articles were identified by the search terms "bone quality" (1851 articles), "anabolic agent" (5044 articles), "PTH or parathyroid hormone" (32,229 articles), "strontium" or "strontium ranelate" (283 articles), "prostaglandin" (77,539 articles), and "statin" or "statins" (14,233 articles). The search strategy included combining each with the phrase "bone quality." Another more limited search aimed at finding more novel potential agents. RESULTS: Parathyroid hormone is the only US Food and Drug Administration-approved bone anabolic agent in the United States and has been the most extensively studied in in vitro animal and human trials. Strontium ranelate is approved in Europe but has not undergone Food and Drug Administration trials in the United States. All the studies on prostaglandin agonists have used in vivo animal models and there are no human trials examining prostaglandin agonist effects. The advantages of statins include the long-established advantages and safety profile, but they are limited by their bioavailability in bone. Other potential pathways include proline-rich tyrosine kinase 2 (PYK2) and sclerostin (SOST) inhibition, among others. CONCLUSIONS: The ongoing research to enhance the anabolic potential of current agents, identify new agents, and develop better delivery systems will greatly enhance the management of bone quality-related injuries and diseases in the future.

Circulating fibroblast growth factor-23 increases following intermittent parathyroid hormone (1-34) in postmenopausal osteoporosis: association with biomarker of bone formation.

Uncertainties exist regarding whether FGF-23 production is influenced by PTH and its involvement in bone formation. We evaluated FGF-23 response and its relation to changes in biomarkers of bone formation following intermittent PTH treatment. Twenty-seven women with a mean [SD] age of 75.8 [5.4] years with postmenopausal osteoporosis were treated with PTH(1-34) for 18 months. Bone mineral density (BMD) was measured at 6 and 18 months at the lumbar spine (LS) and total hip (TH). Blood samples were obtained at baseline, 1-3, 6-9, and 12-18 months. Serum calcium, phosphate, PTH, 25(OH)vitamin D, 1,25(OH)(2)vitamin D, markers of bone turnover, FGF-23, and sclerostin were measured. BMD increased at both the LS (11.6%, P < 0.001) and TH (2.5%, P < 0.01). The bone formation marker P1NP increased early (baseline mean [SD] 39.9 [24.4] µg/l, 1-3 months 88 [37.9] µg/l; P < 0.001) and remained higher than baseline throughout 18 months. FGF-23 also increased, with a peak response at 6-9 months (increase 65%, P = 0.002). Serum phosphate remained stable. A significant increase in 1.25(OH)(2)vitamin D (P = 0.02) was seen at 1-3 months only. A small but significant reduction in sclerostin was seen at 6-9 (P = 0.02) and 12-18 months (P = 0.06). There was a positive correlation between changes in P1NP and FGF-23 (6-9 months r = 0.78, P < 0.001). FGF-23 is increased by intermittent PTH(1-34). This is related to early changes in P1NP, suggesting that the skeletal effects of PTH may involve FGF-23. Further studies are required to elucidate this.